1. Field of the Invention
The present invention relates to a method of producing valienamine and, more particularly, to a method of producing valienamine at a substantially high conversion rate. The method involves mass producing valienamine by way of selective hydrolysis by using TFA from acarbose or acarbose derivatives, and then removing its by-products therefrom, i.e., monosaccharides, disaccharides, and trisaccharides.
2. Brief Description of the Related Art
The conventional art relating to commercial production of valienamine can be divided into two types. The first type relates to a method of direction production of valienamine by using microorganism fermentation, and the second type relates to a method of production of validamycin, a derivative of valienamine, by degradation by using other microorganisms.
Validamycin derivatives basically include a valienamine moiety which selectively binds with validamine or valiolamine. Moreover, a validamycin derivative is a pseudotrisaccharide compound which is glucose bonded in chain.
The validamycin compound is an antibiotic used for germicide for rice-cultivated land in East Asia which is produced among other methods by culturing Streptomyces hygroscopicus, a soil microorganism. Here, the validamycin compound contains a small amount of intermediate valienamine which is then separated out via a column.
As for another method for producing valienamine, there is a method of separating validamycin by using a microorganism, F. saccharophilum, etc. The method involves using validamycin as a substrate or medium and adding it to the liquid mixed with microorganisms; culturing them for a certain period of time; and then inducing separation of validamycin by microorganisms; and then obtaining validamycin by separation via a column. Yet, the two methods have disadvantages in that they take too much time for microorganism fermentation with not much higher yield.
Another compound which has a valienamine moiety is acarbose. Acarbose is obtained from secondary metabolic products of Actinoplanes sp., which is one type of soil microorganisms. It is currently being used as a treatment for diabetes, since it has inhibition effects on α-amylase. However, as of yet, there is no disclosure of the process of commercial or mass production of valienamine by using acarbose as a raw material.
As for methods of producing valienamine, reported in academia, there is a chemical method of producing valienamine by using N-bromosuccinimide (NBS) with validamycin as raw material. However, as this method uses dimethyl sulfoxide (DMSO) as solvent, it suffers from difficulties during purification and separation processes of byproducts, in addition to its low yield. Moreover, there is an ongoing research into the production method of valienamine using organic and inorganic acid, such as sulfuric acid, hydrochloric acid, and acetic acid. Yet, the method is not practical since it is limited to the extent of hydrolysis of only one terminal saccharide. Moreover, there was an attempt to produce valienamine by way of organic synthesis, but it currently is in a standstill due to the inefficiency of purification and organic synthesis processes.
There is a production method of valienamine by pre-synthesis by enzyme, which is being actively pursued in recent years. This is a method of producing valienamine by using an inexpensive substrate by finding valienamine synthesis and related enzymes expressed by a strain. However, there are many difficulties, such as determination of the degree of activity and the expression according to a gene probe. So, the current production is rather difficult.
As stated above, the production of valienamine in vitro by purification enzymes or chemicals has not yet been commercialized, and so up to now, valienamine has been mainly synthesized and produced by hydrolyzing validoxylamine and validamycin by using the strains, Pseudomonas denitrificans and Flavobacterium saccharophilum. Japanese Patent No. 57,054,593 discloses a reaction of converting validoxylamine and validamycin by using microorganisms. This is a method of synthesizing and producing valienamine by using Flavobacterium saccharophilum by reacting 1-5 wt. % mixture of validoxylamine and validamycin for about 24 to about 200 hours at reaction conditions of 20-45° C. and pH 5-8.